Since hydroxycarboxylic acids are useful as a raw material for polymers or an intermediate for medicines, a method for effectively producing hydroxycarboxylic acids have been demanded.
As an example, glycolic acid (α-hydroxyacetic acid) can be mentioned. Glycolic acid has been used as a raw material for cleaning agents or cosmetics, but has recently received attention as a raw material for polyglycolic acid which is useful as a gas barrier polymer or a medical polymer. The reason why glycolic acid has received attention as a gas barrier material is that a layer of polyglycolic acid has high oxygen barrier property and performance as a material for packing food or carbonated beverage which can easily spoil in the presence of oxygen.
Glycolic acid of a chemically synthesized product which is currently commercially available contains quite a few impurities, which is a problem when used as a raw material for polymers in view of purity. This is because these impurities inhibit a dehydrating condensation reaction of glycolic acid, and also methoxy acetate which is one of those impurities is a compound suspicious of carcinogenic potential, thus being desirable not to be included in a packing material for food or beverage. Of course, it is technically possible to remove impurities by purification, but such the purified products are actually high in cost and thus are not practical as a raw material for packing at low cost.
In order to avoid the aforementioned problems given in glycolic acid of chemically synthesized products, a production of glycolic acid according to a biomethod employing ethylene glycol as a raw material has been attempted.
In Patent Document 1 and Patent Document 2, there has been disclosed a method for producing glycolic acid by a microorganism, which includes culturing yeast belonging to genus Pichia, genus Rhodotorula, genus Sporobolomyces, genus Kluyveromyces or genus Torulopsis, a strain belonging to genus Nocardia, a strain belonging to genus Rhodococcus, or an Escherichia coli B strain in a culturing medium containing ethylene glycol and separating and collecting glycolic acid from the culturing broth.
Among the methods for producing glycolic acid as described in Examples of Patent Document 1 and Patent Document 2, a method employing Pichia naganishii gives the highest accumulation concentration of glycolic acid, and 35.3 g/L of glycolic acid is obtained by a reaction for 30 hours. In regard to the production of glycolic acid with the use of Pichia naganishii, it has been reported in Non-Patent Document 1 that 105 g/L of glycolic acid can be obtained by a reaction for 120 hours with further improved reaction conditions.
In Patent Document 3, it has been described that it is possible to produce hydroxycarboxylic acids including glycolic acid from a raw material like aliphatic polyhydric alcohols having a hydroxyl group at the end such as ethylene glycol, by using a microorganism in which a gene encoding lactaldehyde reductase and a gene encoding lactaldehyde dehydrogenase are introduced in the form of plasmid so as to impart or enhance an activity of those enzymes, as well as described that an ability to produce glycolic acid is improved by disrupting a gene encoding glycolate oxidase contained in a microorganism so as to inactivate an activity of the enzyme.
In a reaction for producing hydroxycarboxylic acids including glycolic acid by the above-mentioned conventional methods, an amount of microbial cell required for the reaction is large, which thereby causes problems such as an increase in the production cost, contamination by impurities derived from the microbial cells, and requiring so much work and cost for disposing the microbial cells after the production of hydroxycarboxylic acids.
As a biosynthesis pathway of nicotinamide adenine dinucleotide in a microorganism, there are a pathway (de novo pathway) in which a biological synthesis is done via quinolic acid from aspartic acid and a pathway (recycle pathway) in which nicotinamide produced by a metabolization of nicotinamide adenine dinucleotide and so on is recycled. It has been known that the biosynthesis pathways thereof in an Enterobacteriaceae family including genus Escherichia, genus Shigella, genus Salmonella, genus Erwinia, genus Yersinia and genus Photorhabdus is controlled by a protein (hereinafter, referred to as NadR) encoded by nadR (may be referred to as nadI according to literatures) gene. In specific, it has been known that NadR inhibits the expressions of an L-aspartic acid oxidase gene and a quinolinic acid synthetase gene in the de novo pathway, as well as a nicotinic acid phosphoribosyltransferase gene (hereinafter, referred to as pncB) in the recycle pathway.
On the other hand, NadR as a multifunctional protein has important functions as described below in the biosynthesis of nicotinamide adenine dinucleotide as described below. That is, it is clear that NadR also has functions as a transfer of nicotinamide mononucleotide which is a precursor for nicotinamide adenine dinucleotide, as well as a function as nicotinamide mononucleotide adenylyltransferase catalyzing the reaction for producing deamide-nicotinamide adenine dinucleotide which is a precursor for nicotinamide adenine dinucleotide from ATP and nicotinic acid ribonucleotide.
Microorganisms in which nadR gene is disrupted have been already reported in Non-Patent Document 2, but a production of hydroxycarboxylic acids by such the microorganisms has not bee reported.
In Non-Patent Document 3, it has been reported that the content of nicotinamide adenine dinucleotide is enhanced by introducing a pncB expression vector to Escherichia coli.     [Patent Document 1] Japanese Patent Laid-open No. H10-174593    [Patent Document 2] Japanese Patent Laid-open No. H10-174594    [Patent Document 3] International Publication Pamphlet No. WO 2005/106005    [Non-Patent Document 1] Biosci. Biotechnol. Biochem., Vol. 65 (10), pp. 2265-2270, (2001)    [Non-Patent Document 2] J. Bacteriol., Vol. 187 (8), pp. 2774-2784, (2005)    [Non-Patent Document 3] Metabolic Engineering, Vol. 4, pp. 238-247, (2002)